目的: 建立超高效液相色谱串联三重四极杆质谱法(UPLC-MS/MS),在12 min内完成对绞股蓝药材中16个皂苷类成分(人参皂苷CK、F2、Rb1、Rb3、Rd、Rg2、Rg3、Rg5、Rg6、Rh4、Rh8,绞股蓝皂苷A、XLVI、XLIX,三七皂苷Fd和越南人参皂苷R8)的含量测定。方法: 采用UPLC-MS/MS法,正离子多反应监测(MRM)模式进行含量测定。色谱条件:采用Waters CORTECS C18色谱柱(2.1 mm×100 mm, 1.6 μm),流动相为0.1%甲酸水(A)-乙腈(B),梯度洗脱,流速为0.25 mL·min-1,柱温为45 ℃。结果: 在所设定的色谱条件下,12 min内完成对绞股蓝药材中上述16个成分的含量测定,在考察的浓度范围内均呈良好的线性关系(r>0.997 6),回收率和RSD分别在96.2%~104.0%和1.5%~2.9%,精密度、重复性、稳定性考察均符合分析要求。含量测定结果发现,16个皂苷中,绞股蓝皂苷XLVI、A、XLIX和人参皂苷Rb3含量较高,含量均值分别达到7.083、5.599、1.657、1.283 mg·g-1,且不同产地的绞股蓝药材中皂苷类成分存在较大差异,聚类分析可分为Ⅰ、Ⅱ、Ⅲ 3类。统计发现,越南人参皂苷R8,人参皂苷Rb1、Rb3、Rd、Rg3、Rg5、Rg6、Rh8、F2、CK,绞股蓝皂苷XLVI和三七皂苷Fd成分含量在类型Ⅱ中高于类型Ⅰ和Ⅲ,而绞股蓝皂苷A与XLIX成分含量在Ⅲ类型中高于类型Ⅰ和Ⅱ。结论: 本研究所建立的在12 min内完成对绞股蓝中16个皂苷类成分的UPLC-MS/MS定量分析方法简便、快捷、准确,分离效果较好,可为完善绞股蓝中皂苷类成分的质量标准评价提供依据。
Objective: To establish an UPLC-MS/MS method for simultaneous determination of 16 major components (ginsenoside Rh4, ginsenoside Rh8, ginsenoside Rg6, ginsenoside Rg5, ginsenoside F2, ginsenoside Rd, ginsenoside Rb3, ginsenoside Rb1, ginsenoside C-K, ginsenoside Rg3, ginsenoside Rg2, gypenoside A, gypenoside XLVI, gypenoside XLIX, notoginsenoside Fd and vina-ginsenoside R8) in Gynostemmatis Pentaphylli Herba. Methods: The UPLC-MS/MS assay was performed on a Waters CORTECS C18 (2.1 mm×100 mm,1.6 μm) column with acetonitrile-water (containing 0.1% formic acid) as mobile phase by gradient elution at a flow rate of 0.25 mL·min-1. The column temperature was set at 45 ℃. MS detection was performed with multiple reaction monitoring (MRM) mode using positive electrospray ionization. Results: Under the optimized chromatographic conditions, good separation of 16 saponins were obtained within 12 min. Good linearities were found in the concentration ranges (r>0.997 6), the recoveries and RSDs were in the range of 96.2%-104.0% and 1.5%-2.9%respectively. And the precision, repeatability and stability also met the analysis requirements. The contents of gypenoside XLVI, gypenoside A, gypenoside XLIX and ginsenoside Rb3 were higher, with the average contents of 7.083, 5.599, 1.657 and 1.283 mg·g-1, respectively. There were significant differences in the contents of gypenoside in samples from different habitats. Through cluster analysis, they could be divided into three categories. The contents of ginsenoside R8, ginsenoside Rb1, ginsenoside Rb3, gynostemma gynostemma ginsenoside XLVI, ginsenoside Rh8, ginsenoside Rd, ginsenoside Fd, ginsenoside Rg6, ginsenoside F2, ginsenoside Rg3, ginsenoside CK and ginsenoside Rg5 in type Ⅱ were higher than those in type Ⅰ and Ⅲ. And the contents of gypenosides A and XLIX in type Ⅲ were higher than those in type Ⅰ and Ⅱ. Conclusion: It is the first report about simultaneous analysis of 16 saponins in Gynostemmatis Pentaphylli Herba by using UPLC-MS/MS method, which affords highly sensitive, specific, fast and efficient method for quality control of Gynostemmatis Pentaphylli Herba.
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